Thai J Toxicology 2008; 23(2):135-142 P09/ 135

Antimutagenicity of Some Thai Dishes on Urethane Induced Somatic Mutation and Recombination in Drosophila melanogaster

Kangsadalampai K* and Pratheepachitti N

Institute of Nutrition, Mahidol University, Salaya, Nakhon Pathom 73170,

ABSTRACT

This study examined the mutagenicity of Thai dishes, namely Thai main dishes (Tom Yam Kung, Kaeng Liang, Kaeng Som Pak Ruam, NamPrik Kapi, Nam Prik Makam, and Yam Tua Pu) and Thai one dish meals (Khaow Yam Pak Tai, Khanomjeen Nam-ngiew and Khaow Man Som Tam). The antimutagenicity of the samples on urethane (URE) induced somatic mutation and recombination in Drosophila melanogaster was also determined. Eighty trans-heterozygous Drosophila melanogaster larvae, aged three-days old, obtained from virgin ORR; flr3 virgin female and mwh male were transferred to a test tube containing each Thai dish mix with regular medium (mutagenicity study) or regular medium containing 36 mM URE (antimutagenicity study) until they became adult flies. The ratios (w/w) of Thai dish and a mixture of regular medium or regular medium containing URE were 1:1, 1:2 and 1:4. The occurrences of mutant spots on the round wing of surviving flies were analyzed. It was found that all Thai dishes were not mutagenic. The antimutagenicity of three kinds of Thai dishes at ratios of 1:1 and 1:2 were 61-94 percent inhibition and at a ratio of 1:4 were about 45 – 83 percent inhibition. The antimutagenic mechanisms were not clearly elucidated in this study but rather suggested the effects of many antimutagens in the components of each dish. The findings from the present experiment seems to justify the claim that Thai dished are good for health, aside from its superb sensory attributes as produced by mixtures of different ingredients.

Keywords: Mutagenicity, antimutagenicity, Thai dishes, urethane, SMART

*Corresponding author: Dr. Kaew Kangsadalampai Institute of Nutrition, Mahidol University, Salaya Nakhonpathom, 73170, Thailand E-mail: [email protected]

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Introduction Many traditional cuisines, such as Mediterranean cookery, Japanese food preparation, and Thai diet are interesting because of their health benefits. Thai traditional diet is characterized with high amount of vegetables, fruits, herbs and spices.1 Thai people consume in large quantity mixtures of various kinds of spices and prepared as pastes. Several studies reported that components of diet could be a major factor in modulating the risk of cancer, for instance, Thai edible plants have been reported for their antimutagenic or anticarcinogenic potency, in vitro and in vivo.2-5 However, most studies used the extract or the unprocessed plants rather than the dishes of complex mixtures of many ingredients that may interact with each other. Only curry pastes which are mixtures of dried chilies, , and other ingredients depending on types of curry paste have been evaluated for possible antigenotoxicity.6 They are used as the main ingredient in dishes but no in-vivo study to evaluate the antimutagenic effect of Thai dishes was undertaken. Therefore, the Drosophila somatic mutation and recombination test (SMART) system has been employed in the present study to assess the effect of various Thai dishes in modulating the genotoxicity of urethane which is a promutagen metabolically activated by the cytochrome P-450 enzyme system.7

Materials and Methods Chemicals and Samples: Urethane (URE) was purchased from Sigma Chemical (St. Louis, Mo, USA). Food Chemistry Division (Institute of Nutrition Mahidol University) provided the dishes for this experiment. Ingredients of each Thai dish are shown in Table 1. Each sample was homogenized as paste and kept refrigerated until used. Other chemicals were of laboratory grade. Experimental Design Virgin females of Oregon wing flare strain (ORR/ORR; flr3/TM3, Ser) were mated with males of multiple wing hair strain (mwh/mwh) on regular medium to produce trans-heterozygous larvae of improved high bioactivation cross (IHB). Both strains were obtained from the Institute of Toxicology (Swiss Federal Institute of Technology, and the University of Zurich) and maintained on the regular medium modified from the formula of Roberts8 which had propionic acid (0.01 ml) as a preservative. Appropriate amount of each Thai dish was added to regular medium at the ratio of 1:1, 1:2, or 1:4 w/w and it was homogenized; the final percentage of sample in each experimental media was 50%, 33% or 25%, respectively. Equal amount of each mixed medium was transferred into a 15 ml test tube. Each medium was used as an experimental medium for mutagenicity testing of each dish. URE (36 mM) was substituted for deionized water in the regular medium and was used as a positive control medium. An experimental medium containing URE was prepared by adding each Thai dish into the positive control medium at the same ratio described above and homogenized. Equal amount of each mixed medium was transferred into a 15 ml test tube. This medium was used for antimutagenicity study. The mutagenicity of each sample (in the experimental medium) was assayed as described by Graf et al.9 and the antimutagenicity of each sample was assayed using the experimental medium containing URE. The larvae were maintained on medium at 25+1oC until pupation. The surviving adult flies bearing the marker trans-heterozygous (mwh+/+flr3) indicated with round wings were collected. Subsequently, the wings were removed, mounted and scored under a compound microscope for recording of the wing spot. Induction frequencies of wing spots of Thai dishes treated groups were compared with that of the deionized water negative control group. The estimation of spot frequencies and confidence limits of the estimated mutation frequency were performed with significant level of α = β = 0.05. A multiple-decision procedure was used to decide whether a sample was positive, weak positive, inconclusive or negative mutagen as described by Frei and Wurgler.10 Antimutagenicity was estimated using percentage of inhibition of total spots per wing calculated as follows: percentage of inhibition = (a-b)/a x 100. Where “a” is the number of total spots per wing induced by 1st National Conference in Toxicology 17-18 November 2008 Thai J Toxicology 2008 137

URE, “b” is the number of total spots per wing induced with URE administered with each Thai dish. It was proposed that percent of inhibition between 0–20%, 20–40%, 40–60% and higher than 60% were classified as negligible, weak, moderate and strong antimutagenicity, respectively.

Results Table 1 presents the common ingredients and amount of each recipe. All Thai dishes, namely Thai main dishes (Table 2) and Thai one-dish meals (Table 3) reduced the number of URE-induced wing spots when each dish, along with URE, was administered to the three-days-old larvae. Most dishes added to the positive control medium at the ratio 1:1 and 1:2 showed strong antimutagenicity against the genotoxicity of URE (61-94% inhibition). Only the ratio 1:4 of some sample to the positive control medium revealed moderate antimutagenicity. Similar trends were obtained in both first and second trials. Only Kanomjeen Nam-ngiew (Table 3) showed weak to strong antimutagenicity effect against URE depending on the amount of the dish in the medium. This revealed that percentage of inhibition is dependent on the amount of each Thai dish added to the fly medium.

Discussion Safety of Thai Dishes: Traditional Thai dishes are safe in terms of mutagenicity as resulted from Drosophila melanogaster tests. The average size and survival rates of adult flies obtained from larvae fed on medium containing each Thai dish with 1:1 ratio did not show any difference compared with the control group (fed on regular medium). Only the larvae fed on the highest amount (1:1 ratio) of either Nam Prik Makam or Nam Prik Kapi had smaller size and lower number of surviving adult flies. These dishes that contain table salt might retard the growth of larvae or even killed the larvae. Analysis performed by the Division of Food Chemistry, Institute of Nutrition, showed that both Nam Prik Kapi and the Nam Prik Makam contained 22 mg sodium per 100 g. Kangsadalampai and Sommani11 found that size of Drosophila larvae fed on salty fermented soybean products namely; soy paste (26 mg sodium chloride per g) and sufu which was preserved bean curd (37 mg sodium chloride per g) were smaller than that of the negative control group and also had lower survival rates. However, this should not pose any problem to consumer since both dishes have strong flavor (i.e. hot, salty, sweet and sour) because only small amount is consumed with large amount of fresh, steamed or boiled vegetables or deep-fried mackerels. The high unsaturated fatty acids content of vegetable oil used in Nam Prik Makam could contribute to high level of free radicals that may cause toxicity on Drosophila melanogaster. This organism generally lacks superoxide dismutase,12,13 thus, some have no resistance to its toxic content. To prevent this effect of experimental medium to fly development, the amount of sample incorporated in the regular medium was reduced to 1:2 and 1:4. Antimutagenicity of Thai dishes: URE is metabolically activated by cytochrome P-450 enzyme system (7). Vinyl epoxide, the reactive intermediate of URE metabolism, is the carcinogenic active metabolite.14 Kemper15 reported the carcinogenic metabolites of URE are detoxified with glutathione-S-transferase (GST) conjugation. Substantial information indicated that the mutagenicity of URE decreased in the presence of antimutagens or anticarcinogens in many food and beverages. Overall results of the present investigation showed that most Thai dishes could reduce the mutagenicity of URE. This protective outcome could be the result of more than one mechanisms and the antimutagenicity could be the total effect of all ingredients in each Thai dish.

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Table 1 Ingredients of each Thai dish Ingredients of each Thai dish Dish English name Sub-group Food item and amount per recipe (g) Kaeng Liang Thai style main peeled pumpkin (121), hairy basil leaves (73), ivy vegetable gourd leaves (81), sponge gourd (109), mushrooms- soup straw (150), bottle gourd (100), soup stock (1188), chili paste pepper (3.5), peeled , sliced (89), paste (20), ground (44) seasoning (17) Kaeng Som Pak Sour and main snake head fish (215), meat snake-head fish (110), Ruam spicy curry water (1069), long beans (210), young water melon (240), cabbage (230), sesbania flowers (150) Nam Prik Kapi* Dried shrimp main grilled (43), hot chili (4.6), peeled garlic paste dip (18.9), ground dried shrimp (3.5), pea aubergine (36), ripe ma-euk, sliced (20), old round aubergine seed (4), red hot chili (1) seasoning fish sauce (28), lime juice (50), palm sugar (47) Nam Prik dip main peeled young tamarind (95), hot chili (2.8), chopped Makam* peeled garlic (24), grilled shrimp paste (28), soybean oil (23), ground dried shrimp (12) Tom Yam Kung Sour and main giant fresh water (441), mushroom-straw (420), spicy prawn young galangal (15.4), leaves (1.5), lemon soup grass (29), hot chili (4.7), coriander leaves (7), soup (832) seasoning fish sauce (73.6), lime juice (73.8) Khanom-jeen spicy Main Thai noodle or Khanom Jeen (400), small cubes pork Nam-ngiew noodles blood (138), pork chop (75), small tomato (139.4), dried red cotton flowers (2.3), pork cartilage stock (600), chopped pork cartilage (262), water (865) chili paste dried chili (11.5), shrimp paste (3.7), sliced shallots (44.4), sliced peeled garlic (10), sliced galangal (2.8), sliced coriander rhizomes (2.9), dried fermented soybean (22.5), vegetable oil (16.8), fish sauce (62) side dish mung bean sprouts (200), sliced fermented chinese (vegetables) cabbage (80), spring onion, sliced (12), sliced coriander leaves, (4), lime juice (26.8), fried garlic (7.4), fried-dried chili (5.8) Khaow Man Oily Rice With Khaow-Man rice (250), (356), grated coconut (300), Som Tam Spicy water (238), sugar (16), salt (4), pandatus leaves (2 Salad leaves) Som Tam raw papaya (227), peeled garlic (5), dried chili (3), pepper (0.1), tamarind extract (31.5), fish sauce (46.5), palm sugar (62.8), lime juice (29.5), ground dried shrimp (10), hot chili (2.2), small lime peels (8), vegetables lettuce (75), coral leaves (20) Khaow Yam Pak Rice salad main (780), fried sun-dried rice (167), ground Tai dried shrimp (73), roasted grated coconut (96), lime juice (96), ground chili (8.3),pounded (250), salty budu (125), palm sugar (190), pounded lemon grass (30), pounded galangal (18), kaffir lime leaves (3.7), pounded shallot (57.7), water (505) side dish long bean (249), mung bean sprout (395), sliced (vegetable/fruit) cucumber (181), fine sliced kaffir lime leaves (11), fine sliced lemon grass (120), fine sliced wild betel leaves (27), pomelo, edible portion (454) *Generally consumed with a combination of various vegetables

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Table 2 Effect of each Thai main dishes on URE-treated Drosophila melanogaster

% of Spots per wing a (Number of spots from 40 wings) Antimuta- sample Percent Sample Small single Large single Twin Total genicity in the fly Inhibition classification medium (m=2.0) (m=5.0) (m=5.0) (m=2.0) First trial Water - 0.2(8) 0.00(0) 0.00(0) 0.2(8) - - 36 mM Urethane - 13.30(532)+ 2.68(107)+ 0.30(12)+ 16.28(651)+ 0 - Tom Yam Kung 50 2.85(114)+ 0.98(39)+ 0.22(9)+ 4.05(162)+ 76 strong 33 2.95(118)+ 0.60(24)+ 0.10(4)+ 3.65(146)+ 78 strong 25 5.70(228)+ 1.15(46)+ 0.10(4)+ 6.95(278)+ 59 moderate Kaeng Liang 50 1.80(72)+ 0.05(2)+ 0.00(0) 1.85(74)+ 87 strong 33 3.18(127)+ 0.40(16)+ 0.10(2)+ 3.63(145)+ 74 strong 25 5.13(205)+ 0.85(34)+ 0.20(8)+ 6.18(247)+ 56 moderate Kaeng Som Pak Ruam 50 3.40(136)+ 0.97(39)+ 0.08(3)+ 4.45(178)+ 68 strong 33 3.32(133)+ 1.45(58)+ 0.17(7)+ 4.95(198)+ 65 strong 25 4.78(191)+ 2.45(98)+ 0.32(13)+ 7.55(302)+ 46 moderate Nam Prik Kapi 50 3.58(143)+ 0.25(10)+ 0.15(6)+ 3.98(159)+ 78 strong 33 3.60(144)+ 0.55(22)+ 0.23(9)+ 4.38(175)+ 76 strong 25 6.35(254)+ 1.25(50)+ 0.22(9)+ 7.82(313)+ 57 moderate Nam Prik Makam 50 2.56(41)+ 0.38(6)+ 0.00(0) 2.94(47)+ 83 strong 33 6.40(256)+ 2.00(80)+ 0.30(13)+ 8.70(348)+ 49 moderate 25 1.65(66)+ 0.48(19)+ 0.07(3)+ 2.20(88)+ 86 strong Yam Tua Pu 50 3.85(154)+ 0.85(34)+ 0.15(6)+ 4.85(194)+ 71 strong 33 3.70(148)+ 0.70(28)+ 0.40(16)+ 4.80(192)+ 71 strong 25 4.40(176)+ 1.52(61)+ 0.40(16)+ 6.32(253)+ 62 Strong Second trial Water - 0.18(7) 0.00(0) 0.00(0) 0.18(7) - - 36 mM Urethane - 12.77(511)+ 4.00(160)+ 0.28(11)+ 17.05(682)+ 0 - Tom Yam Kung 50 1.88(75)+ 0.32(13)+ 0.15(6)+ 2.35(94)+ 86 strong 33 3.78(151)+ 1.25(50)+ 0.25(10)+ 5.28(211)+ 69 strong 25 4.85(194)+ 1.35(54)+ 0.15(6)+ 6.35(254)+ 63 Strong Kaeng Liang 50 1.33(53)+ 0.02(1)+ 0.00(0) 1.35(54)+ 91 strong 33 1.82(73)+ 0.43(17)+ 0.05(2)+ 2.30(92)+ 85 strong 25 3.00(120)+ 0.92(37)+ 0.08(3)+ 4.00(160)+ 73 Strong Kaeng Som Pak Ruam 50 3.10(124)+ 0.90(36)+ 0.28(11)+ 4.28(171)+ 71 strong 33 3.83(153)+ 1.40(56)+ 0.32(13)+ 5.55(222)+ 63 strong 25 4.88(195)+ 2.52(101)+ 0.28(11)+ 7.68(307)+ 49 moderate Nam Prik Kapi 50 3.48(139)+ 0.60(24)+ 0.10(4)+ 4.18(167)+ 76 strong 33 4.15(166)+ 0.95(38)+ 0.20(8)+ 5.30(212)+ 70 strong 25 6.52(261)+ 0.92(37)+ 0.28(11)+ 7.72(309)+ 56 moderate Nam Prik Makam 50 4.15(83)+ 0.45(9)+ 0.30(6)+ 4.90(98)+ 71 strong 33 4.18(167)+ 1.60(64)+ 0.22(9)+ 6.00(240)+ 65 strong 25 4.92(197)+ 0.93(37)+ 0.35(14)+ 6.20(248)+ 63 strong Yam Tua Pu 50 4.78(191)+ 0.90(36)+ 0.27(11)+ 5.95(238)+ 65 strong 33 4.12(165)+ 0.75(30)+ 0.48(19)+ 5.35(214)+ 69 strong 25 4.88(195)+ 1.90(76)+ 0.27(11)+ 7.05(282)+ 59 moderate a Statistical diagnoses using estimation of spot frequencies and confidence limits according to Frei and Wurgler (1988) for comparison with deionized water : + = positive, - = negative; m = multiplication factor. Probability levels: α = β = 0.05. Using one-sided statistical tests.

The modulation detoxifying system could be a mechanism to inhibit the mutagenicity of URE. Citrus plants used in Thai dishes, namely, lemon grass, kaffir lime leaves and lime juice contain some bitter compounds e.g., limonene, naringenin, naringin, diosmin, tangeretin and rutin. Many citrus flavonoids (phenolic compounds) have been reported for their antimutagenicity against many mutagens by modulating the detoxifying enzymes of the host.16,17 In this study, garlic and shallot, the most common herbal

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Table 3. Effect of each Thai one-dish meal on URE-treated Drosophila melanogaster a Percent of Spots per wing (Number of spots from 40 wings) Antimuta- Sample sample in Small single Large single Twin Total Percent genicity the fly m=2.0 m=5.0 m=5.0 m=2.0 inhibition classification medium First trial Water - 0.13(5) 0.02(1) 0.00(0) 0.15(6) - - Urethane - 10.92(437)+ 2.88(115)+ 0.27(11)+ 14.07(563)+ 0 - Khaow Yam Pak Tai 50 0.90(36)+ 0.08(3)+ 0.00(0) 0.98(39)+ 93 strong 33 1.68(67)+ 0.18(7)+ 0.02(1)+ 1.88(75)+ 87 strong 25 2.40(96)+ 0.35(14)+ 0.10(4)+ 2.85(114)+ 80 strong Kanomjeen Nam- 50 3.02(121)+ 1.08(43)+ 0.25(10)+ 4.35(174)+ 74 strong ngiew 33 5.72(229)+ 1.70(68)+ 0.33(13)+ 7.75(310)+ 54 moderate 25 4.20(168)+ 1.9(76)+ 0.15(6)+ 6.25(250)+ 63 strong Khaow Man Som 50 1.65(66)+ 0.40(16)+ 0.12(5)+ 2.17(87)+ 87 strong Tam 33 3.80(152)+ 1.48(59)+ 0.20(8)+ 5.48(219)+ 66 strong 25 3.75(150)+ 1.85(74)+ 0.15(6)+ 5.75(230)+ 65 strong Second trial Water - 0.18(7) 0.00(0) 0.00(0) 0.18(7) - - Urethane - 11.72(469)+ 4.78(191)+ 0.28(11)+ 16.78(671)+ 0 - Khaow Yam Pak 50 0.95(38)+ 0.05(2)+ 0.00(0) 1.00(40)+ 94 strong Tai 33 1.78(71)+ 0.02(1)+ 0.05(2)+ 1.85(74)+ 89 strong 25 2.55(102)+ 0.35(14)+ 0.00(0)+ 2.90(116)+ 83 strong Kanomjeen Nam- 50 2.52(101)+ 0.68(27)+ 0.12(5)+ 3.32(133)+ 81 strong ngiew 33 4.00(160)+ 1.75(70)+ 0.20(8)+ 5.95(238)+ 65 strong 25 7.03(281)+ 3.40(136)+ 0.25(10)+ 10.68(427)+ 38 weak Khaow Man Som 50 3.37(135)+ 1.00(40)+ 0.25(10)+ 4.62(185)+ 70 strong Tam 33 4.30(172)+ 1.48(59)+ 0.20(8)+ 5.98(239)+ 61 strong 25 4.07(163)+ 1.30(52)+ 0.28(11)+ 5.65(226)+ 63 strong a Statistical diagnoses using estimation of spot frequencies and confidence limits according to Frei and Wurgler (1988) for comparison with deionized water : + = positive, - = negative; m = multiplication factor. Probability levels: α = β = 0.05. Using one-sided statistical tests. ingredients in Thai dishes were used. Many organosulfur compounds such as diallyl sulfide (DAS) and diallyl disulfides (DADS) could increase the expression of glutathione-S-transferase (GST) in red blood cells of rats.18 These compounds modulated levels of cytochrome P450 isozymes and increased activity of epoxide 19,20 hydrolase and glutathione-S-transferase and reduced the genotoxicity of aflatoxin B1 and N- nitrosodimethylamine (NDMA) in rat.21 Curry pastes commonly consumed in Thailand contain garlic and shallot as major ingredients, showed antimutagenicity against URE in Drosophila melanogaster.6 Many carotenoids found in ivy gourd, pumpkin, pepper and hairy basil showed their antimutagenic activities in many studies.22,23 Carotenoids are known antioxidants both in vitro24 and in vivo15, therefore, they can counteract some mutagens that require metabolic activation through cytochrome P-450 system25,26 to oxidise them to ultimate mutagens. Further studies to explain the antimutagenic mechanism of carotenoids are still necessary. Moreover, some components that may be present in Thai dishes such as organosulfur compounds and flavonoids inhibited DNA-adduct by scavenging the reactive species of the mutagen.27,28 Since the study was conducted as the co-administration of URE with each dish, desmutagenic activity of some components may interfere with the availability of URE in young larvae. Many vegetables, herbs and spices contain dietary fiber and chlorophyll. Antimutagenicity was observed when some dietary

1st National Conference in Toxicology 17-18 November 2008 Thai J Toxicology 2008 141 fiber such as lignin and suberin adsorb the mutagens29-31 and chlorophyll formed complex with mutagens.32 In vitro or in vivo studies on free radical scavenging activities of dietary fiber such as pectin on colon mucosa of rats were reported. Alkali-lignin inhibited both enzymatic and non-enzymatic lipid peroxidation on cell culture. Lignin and ferulic acid in wheat bran acted as a nitrite scavenger on cell culture.33-35 However, there has been no information on the scavenging activity of these compounds on URE; thus, further investigations would be relevant. It seems to justify the claim that Thai dishes are good for health, aside from its superb sensory attributes as produced by mixtures of different ingredients. The protective effects of each dish may be due to the presence of antimutagenic ingredients. However, this study investigated only the result of co- administration of various Thai dishes with URE. The level of protection may be clearer when the experiments are extended to be pre-feeding study.

References 1. Maruekin P. Development of reference recipes for commonly consumed Thai side dishes and their nutritive values. [MS Thesis in Nutrition]. Bangkok: Faculty of Medicine, Mahidol University; 2001, p 11. 2. Kusamran WR, Ratanavila A, Tepsuwan A. Effects of neem flowers, Thai and Chinese bitter gourd fruits and sweet basil leaves on hepatic monooxygenases and glutathione-S-transferase activities, and in vitro metabolic activation of chemical carcinogens in rats. Food Chem Toxicol 1998;36:475-84. 3. Kusamran WR, Tepsuwan A, Kupradinun P. Antimutagenic and anticarcinogenic potentials of some Thai vegetables. Mutat Res 1998;402:247–58. 4. Chewonarin T, Kinouchi T, Kataoka K, et al. Effects of Roselle (Hibiscus sabdariffa Linn.), a Thai medicinal plant, on the mutagenicity of various known mutagens in Salmonella typhimurium and on formation of aberrant crypt foci induced by the colon carcinogens azoxymethane and 2-amino-1-methyl-6- phenylimidazo[4,5-b]pyridine in F344 rats. Food Chem Toxicol 1999;37:591-601. 5. Tepsuwan A, Kupradinun P, Kusamran WR. Effect of Siamese cassia leaves on the activities of chemical carcinogen metabolizing enzymes and on mammary gland carcinogenesis in the rat. Mutat Res 1999;428:363–373. 6. Kangsadalampai K, Laohavechvanich P, Prasarchimontri P. Effect of Thai curry paste on somatic mutation and recombination induced by urethane in Drosophila melanogaster. J Nutr Assoc Thai 2004;39:35-47. 7. Schlatter J, Lutz WK. The carcinogenic potential of ethyl carbamate (urethane): risk assessment at human dietary exposure levels. Food Chem Toxicol 1990;28:205-11. 8. Roberts DB. Basic Drosopila care and techniques. In: Roberts DB, ed. Drosophila: a pratical approach. IRL Press, Oxford; 1986. p. 1-38. 9. Graf U, van Schaik N. Improved high bioactivation cross for the wing somatic mutation and recombination test in Drosophila melanogaster. Mutat Res 1992;271:59-67. 10. Frei H, Wurgler FE. Statistical methods to decide whether mutagenicity test data from Drosophila assay indicate a positive, netative, or inconclusive result. Mutat Res 1988;203:297-308. 11. Kangsadalampai K, Sommani P. Antimutagencity on urethane of various soybean products using in vivo somatic mutation and recombination test. Thai J Pharm Sci 2003;27:17-32. 12. Rogina B, Helfand SL. Cu, Zn superoxide dismutase deficiency accelerates the time course of an age- related marker in Drosophila melanogaster. Biogerontology 2000;1:163-9. 13. Missirlis F, Phillips JP, Jackle H. Cooperative action of antioxidant defense systems in Drosophila. Curr Biol 2001;11:1272-7. 14. Dahl GA, Mille JA, Mille EC. Vinyl carbamate as a promutagen and a more carcinogenic analog of ethyl carbamate. Cancer Res 1978;38:3793-804.

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15. Kempe RA, Myers SR, Hurst HE. Detoxification of vinyl carbamate epoxide by glutathione: evidence for participation of glutathione-S-transferases in metabolism of ethyl carbamate. Toxicol Appl Pharmacol 1995;135:110-8. 16. Higashimoto M, Yamato H, Kinouchi T, et al. Inhibitory effects of citrus fruits on the mutagenicity of 1- methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid treated with nitrite in the presence of ethanol. Mutat Res 1998;415:219-26. 17. Bea Wl, Teel RW. This will give more definite findings that Thai dishes have a chance to increase the protective mechanism in man. Effects of citrus flavonoids on the mutagenicity of heterocyclic amines and on cytochrome P450 1A2 activity. Anticancer 2000;20:3609-14. 18. Wu CC, Sheen LY, Chen HW, et al. Effects of organosulfur compounds from garlic oil on the antioxidation system in rat liver and red blood cells. Food Chem Toxicol 2001;39:563-9. 19. Guyonnet D, Belloir C, Suschetet M, Siess MH, Le Bon AM. Liver subcellular fractions from rats treated by organosulfur compounds from allium modulate mutagen activation. Mutat Res 2000;466:17-26. 20. Guyonnet D, Belloir C, Suschetet M, et al. Antimutagenic activity of organosulfur compounds from Allium is associated with phase II enzyme induction. Mutat Res 2001;495:135-45. 21. Le Bon AM, Roy C, Dupont C, et al. In vivo antigenotoxic effects of dietary allyl sulfides in the rat. Cancer Lett 1997;114:131-4. 22. Rauscher R, Edenharder R, Platt KL. In vitro antimutagenic and in vivo anticlastogenic effects of carotenoids and solvent extracts from fruits and vegetables rich in carotenoids. Muta Res 1998;413:129-42. 23. De Mejìa EG. Quintanar-Hernandez A, Loarca-Pina G. Antimutagenic activity of carotenoids in green peppers against some nitroarenes. Mutat Res 1998;416:11-9. 24. Sie H, Stahl W, Sundquist AR. Antioxidant functions of vitamins: Vitamin E and C, β-carotene and other carotenoids. Ann NY Acad Sci 1992;669:7-20. 25. Gradelet S, Le Bon AM, Bergis R, et al. Dietary carotenoids inhibit aflatoxin B1-induced liver preneoplastic foci and DNA damage in the rat: Role of the modulation of aflatoxin B1 metabolism. Carcinogenesis 1998;19:403-11. 26. Weisburger JH, Dolan L, Pittman B. Inhibition of PhIP mutagenicity by caffein, lycopene, daidzein and genistein. Mutat Res 1998;416:125-8. 27. Schaffe EM, Liu JZ, Green J, et al. Garlic and associated allyl sulfur components inhibit N-methyl-N- nitrosourea induced mammary carcinogenesis. Cancer Lett 1996;102:199-204. 28. Edenharder R, Sager JW, Glatt H, et al. Protection by beverages,fruits, vegetables, herbs, and flavonoids against genotoxicity of 2-acetylaminofluorene and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in metabolically competent V79 cells. Mutat Res 2002;521:57-72. 29. Kato T, Takahashi S, Kakugawa K. Loss of heterocyclicamine mutagens by insoluble hemicellulose fiber and high molecular weight soluble polyphenolics of coffee. Mutat Res 1991;246: 169-78. 30. Harris PJ, Triggs CM, Roberton AM, et al. The adsorption of heterocyclic aromatic amines by model dietary fibres with contrasting compositions. Chem Biol Interact 1996;100:13-25. 31. Ferguson LR, Harris PJ. Studies on the role of specific dietary fibres in protection against colorectal cancer. Mutat Res 1996;350:173-84. 32. Dashwood R, Guo D. Antimutagenic potency of chlorophyllin in the Salmonella assay and its correlation with binding constants of mutagen-inhibitor complexes. Environ Mol Mutagen 1993;22:164-71. 33. Moller ME, Dahl R, Bockman OC. A possible role of the dietary fibre product, wheat bran, as a nitrite scavenger. Food Chem Toxicol 1988;26:841-845. 34. Erhardt JG, Lim SS, Bode JC, et al. A diet rich in fat and poor in dietary fiber increases the in vitro formation of reactive oxygen species in human feces. J Nutr 1997;127:706-9. 35. Tazawa K, Yatuzuka K, Yatuzuka M, et al. Dietary fiber inhibits the incidence of hepatic metastasis with the anti-oxidant activity and portal scavenging functions. Hum Cell 1999;12:189-96.

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